The present invention relates to surfactant molecules, including polypeptides, proteins, and a variety of other organic molecules, which are suitable for use in the treatment of respiratory distress syndrome in infants as well as in adults.
Naturally-occurring pulmonary surfactant is a complex mixture of lipids and proteins that promotes the formation of a monolayer at the alveolar air-water interface and, by reducing the surface tension, prevents collapse of the alveolus during expiration. Premature infants, and occasionally full term neonates, may lack sufficient endogenous surfactant for normal lung function. This can give rise to a condition termed respiratory distress syndrome (RDS) which may necessitate mechanical ventilation and administration of hyperbaric oxygen. Such intervention, unfortunately, can produce permanent damage to lung tissue and may cause retinopathy of prematurity (ROP) leading to blindness.
Pulmonary surfactant (PS) lines the alveolar epithelium of mature mammalian lungs. Natural PS has been described as a xe2x80x9clipoprotein complexxe2x80x9d because it contains both phospholipids and apoproteins that interact to reduce surface tension at the lung air-liquid interface. Natural surfactant contains several lipid species of which dipalmitoyl phosphatidylcholine (DPPC) is the major component together with phosphatidylglycerol (PG) and palmitic acid (PA). At least three specific proteins are also associated, termed SP-A, SP-B and SP-C. Of these three, SP-B and SP-C are distinct, low molecular weight, relatively hydrophobic proteins that have been shown to enhance the surface-active properties of surfactant phospholipid mixtures. It is believed that they facilitate transfer of lipids from the bulk phase lamellar organization to the air-water interface and also stabilize the lipid monolayer during expiration. The structure of SP-B (which is alternatively referred to as SP18) is unusual in that charged amino acids (predominantly basic) are located at fairly regular intervals within stretches of otherwise hydrophobic residues. For the domain consisting of residues 59-80 of the native SP-B sequence, these charged groups have been shown to be necessary for biological activity. In addition, natural and synthetic peptides which are modeled on this hydrophobic-hydrophilic domain when combined with DPPC and PG exhibit good surfactant activity.
Surfactant is stored in lung epithelial cells in the form of lamellar bodies and, following export, it undergoes a structural transition to form tubular myelin before giving rise to a monolayer at the air-water interface. It has been proposed that surfactant proteins SP-A, -B and -C may facilitate these structural transitions and stabilize the lipid monolayer during expansion and contraction of the alveolus; however, an understanding of lipid-protein interactions at the molecular level is presently lacking. The present invention, therefore, has important implications not only with respect to the treatment of RDS in infants as well as adults, but also because of the insight it may provide into lipid-protein interactions in general.
Several exogenous surfactant formulations are currently used in the treatment of infant RDS. While these have reduced morbidity and mortality, continual improvements are needed. In particular, because of the complications that can arise due to mechanical ventilation and administration of hyperbaric oxygen, the sooner normal lung function can be established in a premature infant the more favorable will be the clinical outcome.
Consistent with the foregoing, important characteristics in an exogenous surfactant include the ability to spread rapidly to the alveoli following administration and the ability to maintain a stable monolayer at the alveolar air-water interface so that repeated treatment was not required. The within-disclosed compounds and compositions are believed useful in the preparation of superior exogenous surfactants.
The present invention discloses a wide variety of surfactant molecules which may be formulated, prepared and utilized as disclosed herein. In various preferred embodiments of the present invention, it is contemplated that the surfactant molecules comprise dipeptides, larger polypeptides, or proteins. In other preferred embodiments, surfactant molecules comprise a variety of organic molecules, including L-amino acids, D-amino acids, substituted amino acids (e.g., amino acids with modified R groups), amino acid metabolites and catabolites, molecules with xe2x80x9cdesignedxe2x80x9d side chains, and amino acid mimics or analogs. Molecules comprising dipeptides or polypeptides joined by linkages other than peptide bonds are also encompassed by the present invention; indeed, any organic molecule possessing or exhibiting surfactant activity as described herein is a xe2x80x9csurfactant moleculexe2x80x9d as contemplated by the present invention.
Therefore, in various preferred embodiments of the present invention, a wide variety of surfactant polypeptides is disclosed. In one embodiment, a preferred polypeptide comprises at least about 4, and more preferably at least about 10, amino acid residues and no more than about 60 amino acid residues and is constituted by alternating groupings of charged amino acid residues and uncharged amino acid residues such that the polypeptide, when admixed with a pharmaceutically acceptable phospholipid, forms a pulmonary surfactant having a surfactant activity greater than the surfactant activity of the phospholipid alone.
In one preferred embodiment, a surfactant polypeptide comprises at least 10 amino acid residues and no more than about 60 amino acid residues and is constituted by alternating groupings of charged amino acid residues and uncharged amino acid residues as represented by the formula [(Charged)a(Uncharged)b]c(Charged)d, wherein a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3. It is further preferred that the polypeptide, when admixed with a pharmaceutically acceptable phospholipid, forms a pulmonary surfactant having a surfactant activity greater than the surfactant activity of the phospholipid alone.
In another preferred embodiment, the present invention discloses polypeptides including a sequence having alternating groupings of amino acid residues as represented by the formula (ZaJb)cZd, wherein Z is an amino acid residue independently selected from the group consisting of R, D, E, and K; J is an xcex1-aminoaliphatic carboxylic acid; a has an average value of about 1to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
In yet another preferred embodiment, surfactant polypeptides are disclosed which have alternating groupings of amino acids residue regions as represented by the formula (BaUb)cBd, wherein B is an amino acid residue independently selected from the group consisting of H, 5-hydroxylysine, 4-hydroxyproline, and 3-hydroxyproline; U is an amino acid residue independently selected from the group consisting of V, I, L, C, Y, and F; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3. In another preferred variation, B is an amino acid derived from collagen and is preferably selected from the group consisting of 5-hydroxylysine, 4-hydroxyproline, and 3-hydroxyproline.
Another preferred embodiment of the present invention discloses polypeptides including a sequence having alternating groupings of amino acid residues as represented by the formula (BaJb)cBd, wherein B is an amino acid residue independently selected from the group consisting of H, 5-hydroxylysine, 4-hydroxyproline, and 3-hydroxyproline; J is an xcex1-aminoaliphatic carboxylic acid; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3. In one preferred variation, J is an xcex1-aminoaliphatic carboxylic acid having four to six carbons, inclusive. In another preferred variation, J is an xcex1-aminoaliphatic carboxylic acid having six or more carbons, inclusive. In yet another variation, J is preferably selected from the group consisting of xcex1-aminobutanoic acid, xcex1-aminopentanoic acid, xcex1-amino-2-methylpropanoic acid, and xcex1-aminohexanoic acid.
A further preferred embodiment of the present invention discloses polypeptides including a sequence having alternating groupings of amino acid residues as represented by the formula (ZaUb)cZd, wherein Z is an amino acid residue independently selected from the group consisting of R, D, E, and K; U is an amino acid residue independently selected from the group consisting of V, I, L, C, Y and F; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
The present invention also contemplates a variety of surfactant compositions, particularly liposomal surfactants. Thus, in one preferred embodiment, the invention discloses a liposomal surfactant composition prepared from a polypeptide comprising about 10 amino acid residues and no more than about 60 amino acid residues and is constituted by alternating groupings of charged amino acid residues and uncharged amino acid residues, and a pharmaceutically acceptable phospholipid, wherein the polypeptide is present in an amount sufficient to increase the surfactant activity of the composition above that of the phospholipid.
In another preferred variation, a surfactant composition of the present invention comprises a surfactant molecule constituted by alternating groupings of charged and uncharged residues; the residues may be amino acids, modified amino acids, amino acid analogs or derivatives, and the like. Molecules having surfactant activity as disclosed herein are especially preferred for use in compositions of the present invention.
In one preferred variation, a surfactant composition of the present invention includes a polypeptide comprising at least 10 amino acid residues and no more than about 60 amino acid residues constituted by alternating groupings of charged amino acid residues and uncharged amino acid residues as represented by the formula [(Charged)a(Uncharged)b]c(Charged)d, wherein a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
In another preferred embodiment, a surfactant composition comprises a polypeptide including a sequence having alternating groupings of amino acid residues as represented by the formula (ZaJb)cZd, wherein Z is an amino acid residue independently selected from the group consisting of R, D, E, and K; J is an xcex1-aminoaliphatic carboxylic acid; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
In yet another preferred embodiment, a surfactant composition comprises a surfactant polypeptide including a sequence having alternating groupings of amino acid residue regions as represented by the formula (BaUb)cBd, wherein B is an amino acid residue independently selected from the group consisting of H, 5-hydroxylysine, 4-hydroxyproline, and 3-hydroxyproline; U is an amino acid residue independently selected from the group consisting of V, I, L, C, Y, and F; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
Another preferred embodiment of the present invention discloses compositions comprising polypeptides including a sequence having alternating groupings of amino acid residues as represented by the formula (BaJb)cBd, wherein B is an amino acid residue independently selected from the group consisting of H, 5-hydroxylysine, 4-hydroxyproline, and 3-hydroxyproline; J is an xcex1-aminoaliphatic carboxylic acid; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3. In one preferred variation, J is an xcex1-aminoaliphatic carboxylic acid having four to six carbons, inclusive. In another variation, J is preferably selected from the group consisting of xcex1-aminobutanoic acid, xcex1-aminopentanoic acid, xcex1-amino-2-methylpropanoic acid, and xcex1-aminohexanoic acid.
A further preferred embodiment of the present invention discloses compositions comprising polypeptides including a sequence having alternating groupings of amino acid residues as represented by the formula (ZaUb)cZd, wherein Z is an amino acid residue independently selected from the group consisting of R, D, E, and K; U is an amino acid residue independently selected from the group consisting of V, I, L, C, Y and F; a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3.
In various preferred embodiments of the present invention, as noted previously, surfactant compositions also comprise one or more phospholipids, wherein the phospholipid is present in the range of about 50-100 weight percent. The polypeptide:phospholipid weight ratio is in the range of about 1:7 to about 1:1,000 in various preferred surfactant compositions of the present invention. Suitable phospholipids are preferably selected from the following group: 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (dipalmitoylphosphatidylcholine, DPPC); phosphatidyl glycerol (PG); and an admixture of DPPC and PG in a weight ratio of about 3:1.
The surfactant compositions (e.g., liposomal surfactants) of the present invention may further comprise palmitic acid, in various preferred embodiments. Preferably, the phospholipid comprises about 50-90 weight percent and the palmitic acid comprises the remaining 10-50 weight percent of the lipid portion of the surfactant. As in other preferred embodiments, the phospholipid may be selected from the group consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (dipalmitoylphosphatidylcholine, DPPC); phosphatidyl glycerol (PG); and an admixture of DPPC and PG. If an admixture of DPPC and PG is selected, it is preferable that DPPC and PG be present in a weight ratio of about 3:1.
The present invention also discloses preferred methods of treating respiratory distress syndrome in patients of any age, including neonates and adults. One such method comprises administering to a patient in need of such treatment a therapeutically effective amount of a surfactant compositionxe2x80x94preferably, a liposomal surfactant compositionxe2x80x94prepared from a polypeptide (or other surfactant molecule) of the present invention and a pharmaceutically acceptable phospholipid, wherein the polypeptide is combined with the phospholipid in an amount sufficient to increase the surfactant activity of the composition above that of the phospholipid. The present invention also discloses a method of treating respiratory distress syndrome wherein the polypeptide is constituted by at about 10-60 amino acid residues and constituted by alternating groupings of charged amino acid residues and uncharged amino acid residues as represented by the formula [(Charged)a(Uncharged)b]c(Charged)d, wherein a has an average value of about 1 to about 5; b has an average value of about 3 to about 20; c is 1 to 10; and d is 0 to 3. In various preferred embodiments, such a polypeptide, when admixed with a pharmaceutically acceptable phospholipid, forms a pulmonary surfactant having a surfactant activity greater than the surfactant activity of the phospholipid alone.
A wide variety of surfactant molecules, proteins, and polypeptides which are preferred for use according to the disclosed methods are described above and in the sections that follow. Other preferred components of surfactant compositions used as disclosed herein include a variety of phospholipids and palmitic acid, as further described herein.